Research Summary

In previous work we have established the elementary steps leading to correct nucleotide incorporation by HIV reverse transcriptase and have quantified the changes in individual kinetic constants occurring during misincorporation. In addition, we have determined the mechanism of action of a class of nonnucleoside inhibitors and characterized changes leading to resistance against these agents. In current work, we are continuing to explore the mechanisms of multiple drug resistance and examine the phenomenon of reciprocal drug resistance. A better understanding of these phenomena at the structural and mechanistic level could lead to the development of better combination therapies in the treatment of AIDS.

The hepatitis C virus (HCV) affects an estimated 4 million Americans and 170 million people worldwide. The current treatment regime using ribavirin and interferon is associated with severe side-effects and is only marginally effective. We have purified the RNA-dependent RNA polymerase in our laboratory and have developed assays for its RNA polymerase activity in vitro. The goal of the research is twofold: 1) to find out what factors are involved in creating the activated polymerase complex of the hepatitis C virus, and 2) to aid in the development of treatments to stop the HCV polymerase from replicating the HCV genome, while minimizing the side-effects.

Several studies point to the likely role of the mitochondrial DNA polymerase in the toxicity of nucleoside analogs used in the treatment of viral infections such as hepatitis and AIDS. We have recently shown that the kinetics of incorporation of nucleoside analogs are correlated with their toxicity, spanning more than seven orders of magnitude.

Rat Kinesin

Microtubule-Dependent Motor ATPase Mechanism and Force Production

We have recently established the pathway of ATP hydrolysis by kinesin, a motor protein involved in fast axonal transport. The protein is a dimer and shows an alternating site ATPase mechanism which is coupled to the movement of the kinesin along the surface of the microtubule. In current work we are examining the kinetic and structural basis for this alternating site activity in order to establish the mechanism of force production. The work serves as an important model for understanding energy transduction in general, and for exploring the family of kinesin-like ATPases which are responsible for a wide range of microtubule-dependent movements in all eukaryotic cells.